Analysis of sub-kilobase chromatin topology reveals nano-scale regulatory interactions with variable dependence on cohesin and CTCF

Enhancers and promoters predominantly interact within large-scale topologically associating domains (TADs), which are formed by loop extrusion mediated by cohesin and CTCF. However, it is unclear whether complex chromatin structures exist at sub-kilobase-scale and to what extent fine-scale regulatory interactions depend on loop extrusion. To address these questions, we present an MNase-based chromosome conformation capture (3C) approach, which has enabled us to generate the most detailed local interaction data to date and precisely investigate the effects of cohesin and CTCF depletion on chromatin architecture. Our data reveal that cis-regulatory elements have distinct internal nano-scale structures, within which local insulation is dependent on CTCF, but which are independent of cohesin. In contrast, we find that depletion of cohesin causes a subtle reduction in longer-range enhancer-promoter interactions and that CTCF depletion can cause rewiring of regulatory contacts. Together, our data show that loop extrusion is not essential for enhancer-promoter interactions, but contributes to their robustness and specificity and to precise regulation of gene expression.

Download Full-text

High-resolution mapping of regulatory element interactions and genome architecture using ARC-C

10.1101/467506 ◽

2018 ◽

Author(s):

Ni Huang ◽

Wei Qiang Seow ◽

Julie Ahringer

Keyword(s):

High Resolution ◽

Regulatory Element ◽

Regulatory Elements ◽

Genome Architecture ◽

Chromosome Conformation ◽

Regulatory Interactions ◽

C Elegans ◽

Chromatin Regulators ◽

Accessible Region ◽

Resolution Mapping

AbstractInteractions between cis-regulatory elements such as promoters and enhancers are important for transcription but global identification of these interactions remains a major challenge. Leveraging the chromatin accessiblity of regulatory elements, we developed ARC-C (accessible region chromosome conformation capture), which profiles chromatin regulatory interactions genome-wide at high resolution. Applying ARC-C to C. elegans, we identify ~15,000 significant interactions at 500bp resolution. Regions bound by transcription factors and chromatin regulators such as cohesin and condensin II are enriched for interactions, and we use ARC-C to show that the BLMP-1 transcription factor mediates interactions between its targets. Investigating domain level architecture, we find that C. elegans chromatin domains defined by either active or repressive modifications form topologically associating domains (TADs) and that these domains interact to form A/B (active/inactive) compartment structure. ARC-C is a powerful new tool to interrogate genome architecture and regulatory interactions at high resolution.

Download Full-text

Topologically Associating Domains and Regulatory Landscapes in Development, Evolution and Disease

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.702787 ◽

2021 ◽

Vol 9 ◽

Author(s):

Juan J. Tena ◽

José M. Santos-Pereira

Keyword(s):

Gene Regulation ◽

Evolutionary Conservation ◽

Regulatory Elements ◽

Regulatory Interactions ◽

Pathological Conditions ◽

Topologically Associating Domains ◽

Recent Advances ◽

Development And Differentiation ◽

Regulatory Landscapes ◽

Animal Genomes

Animal genomes are folded in topologically associating domains (TADs) that have been linked to the regulation of the genes they contain by constraining regulatory interactions between cis-regulatory elements and promoters. Therefore, TADs are proposed as structural scaffolds for the establishment of regulatory landscapes (RLs). In this review, we discuss recent advances in the connection between TADs and gene regulation, their relationship with gene RLs and their dynamics during development and differentiation. Moreover, we describe how restructuring TADs may lead to pathological conditions, which explains their high evolutionary conservation, but at the same time it provides a substrate for the emergence of evolutionary innovations that lay at the origin of vertebrates and other phylogenetic clades.

Download Full-text

Understanding 3D Genome Organization and Its Effect on Transcriptional Gene Regulation Under Environmental Stress in Plant: A Chromatin Perspective

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.774719 ◽

2021 ◽

Vol 9 ◽

Author(s):

Suresh Kumar ◽

Simardeep Kaur ◽

Karishma Seem ◽

Santosh Kumar ◽

Trilochan Mohapatra

Keyword(s):

Gene Expression ◽

Environmental Stress ◽

Genome Organization ◽

Regulation Of Gene Expression ◽

Three Dimensional ◽

Regulatory Elements ◽

Chromosome Conformation ◽

3D Genome ◽

Chromatin Architecture ◽

Capture Techniques

The genome of a eukaryotic organism is comprised of a supra-molecular complex of chromatin fibers and intricately folded three-dimensional (3D) structures. Chromosomal interactions and topological changes in response to the developmental and/or environmental stimuli affect gene expression. Chromatin architecture plays important roles in DNA replication, gene expression, and genome integrity. Higher-order chromatin organizations like chromosome territories (CTs), A/B compartments, topologically associating domains (TADs), and chromatin loops vary among cells, tissues, and species depending on the developmental stage and/or environmental conditions (4D genomics). Every chromosome occupies a separate territory in the interphase nucleus and forms the top layer of hierarchical structure (CTs) in most of the eukaryotes. While the A and B compartments are associated with active (euchromatic) and inactive (heterochromatic) chromatin, respectively, having well-defined genomic/epigenomic features, TADs are the structural units of chromatin. Chromatin architecture like TADs as well as the local interactions between promoter and regulatory elements correlates with the chromatin activity, which alters during environmental stresses due to relocalization of the architectural proteins. Moreover, chromatin looping brings the gene and regulatory elements in close proximity for interactions. The intricate relationship between nucleotide sequence and chromatin architecture requires a more comprehensive understanding to unravel the genome organization and genetic plasticity. During the last decade, advances in chromatin conformation capture techniques for unravelling 3D genome organizations have improved our understanding of genome biology. However, the recent advances, such as Hi-C and ChIA-PET, have substantially increased the resolution, throughput as well our interest in analysing genome organizations. The present review provides an overview of the historical and contemporary perspectives of chromosome conformation capture technologies, their applications in functional genomics, and the constraints in predicting 3D genome organization. We also discuss the future perspectives of understanding high-order chromatin organizations in deciphering transcriptional regulation of gene expression under environmental stress (4D genomics). These might help design the climate-smart crop to meet the ever-growing demands of food, feed, and fodder.

Download Full-text

Estimating DNA-DNA interaction frequency from Hi-C data at restriction-fragment resolution

10.1101/377523 ◽

2018 ◽

Cited By ~ 2

Author(s):

Christopher JF Cameron ◽

Josée Dostie ◽

Mathieu Blanchette

Keyword(s):

Restriction Fragment ◽

Cross Validation ◽

Three Dimensional ◽

Dna Interaction ◽

Interaction Frequency ◽

Chromosome Conformation ◽

Restriction Fragments ◽

Regulatory Interactions ◽

Topologically Associating Domains ◽

Validation Experiments

AbstractHi-C is a popular technique to map three-dimensional chromosome conformation. In principle, Hi-C’s resolution is only limited by the size of restriction fragments. However, insufficient sequencing depth forces researchers to artificially reduce the resolution of Hi-C matrices at a loss of biological interpretability. We present the Hi-C Interaction Frequency Inference (HIFI) algorithms that accurately estimate restriction-fragment resolution Hi-C matrices by exploiting dependencies between neighboring fragments. Cross-validation experiments and comparisons to 5C data and known regulatory interactions demonstrate HIFI’s superiority to existing approaches. In addition, HIFI’s restriction-fragment resolution reveals a new role for active regulatory regions in structuring topologically associating domains.Availability: https://github.com/BlanchetteLab/HIFI

Download Full-text

Large-Scale Genomic Reorganizations of Topological Domains (TADs) at the HoxD Locus

10.1101/116152 ◽

2017 ◽

Author(s):

Pierre J. Fabre ◽

Marion Leleu ◽

Benjamin H. Mormann ◽

Lucille Delisle ◽

Daan Noordermeer ◽

...

Keyword(s):

Long Range ◽

Transcriptional Activation ◽

Limb Development ◽

Large Scale ◽

Regulatory Elements ◽

Chromatin Domain ◽

Sequence Specificity ◽

Chromosome Conformation ◽

Topological Domains ◽

Genomic Deletions

ABSTRACTBackgroundThe transcriptional activation of Hoxd genes during mammalian limb development involves dynamic interactions with the two Topologically Associating Domains (TADs) flanking the HoxD cluster. In particular, the activation of the most posterior Hoxd genes in developing digits is controlled by regulatory elements located in the centromeric TAD (C-DOM) through long-range contacts. To assess the structure-function relationships underlying such interactions, we measured compaction levels and TAD discreteness using a combination of chromosome conformation capture (4C-seq) and DNA FISH.ResultsWe challenged the robustness of the TAD architecture by using a series of genomic deletions and inversions that impact the integrity of this chromatin domain and that remodel the long-range contacts. We report multi-partite associations between Hoxd genes and up to three enhancers and show that breaking the native chromatin topology leads to the remodelling of TAD structure.ConclusionsOur results reveal that the re-composition of TADs architectures after severe genomic re-arrangements depends on a boundary-selection mechanism that uses CTCF-mediated gating of long-range contacts in combination with genomic distance and, to a certain extent, sequence specificity.

Download Full-text

Rapid characterization of nano-scale structures in large-scale ultra-precision surfaces

Optics and Lasers in Engineering ◽

10.1016/j.optlaseng.2020.106200 ◽

2020 ◽

Vol 134 ◽

pp. 106200

Author(s):

Wenjun Yang ◽

Xiaojun Liu ◽

Chi Hu ◽

Wenlong Lu ◽

Cheng Chen ◽

...

Keyword(s):

Large Scale ◽

Nano Scale ◽

Ultra Precision ◽

Scale Structures ◽

Rapid Characterization

Download Full-text

Exploring different sequence representations and classification methods for the prediction of nucleosome positioning

10.1101/482612 ◽

2018 ◽

Cited By ~ 1

Author(s):

Nikos Kostagiolas ◽

Nikiforos Pittaras ◽

Christoforos Nikolaou ◽

George Giannakopoulos

Keyword(s):

Gene Expression ◽

Dna Sequences ◽

Large Scale ◽

Genomic Sequence ◽

Critical Role ◽

Regulation Of Gene Expression ◽

Nucleosome Positioning ◽

Regulatory Elements ◽

Machine Learning Algorithms ◽

N Gram

Nucleosomes form the first level of DNA compaction and thus bear a critical role in the overall genome organization. At the same time, they modulate chromatin accessibility and, through a dynamic equilibrium with other DNA-binding proteins, may shape gene expression. A number of large-scale nucleosome positioning maps, obtained for various genomes, has compelled the importance of nucleosomes in the regulation of gene expression and has shown constraints in the relative positions of nucleosomes to be much stronger around regulatory elements (i.e. promoters, splice junctions and enhancers). At the same time, the great majority of nucleosome positions appears to be rather flexible. Various computational methods have in the past been used in order to capture the sequence determinants of nucleosome positioning but, as the extent to which DNA sequence preferences may guide nucleosome occupancy largely varies, this has proved to be rather difficult. In order to focus on highly specific sequence attributes, in this work we have analyzed two well-defined sets of nucleosome-occupied sites (NOS) and nucleosome-free-regions (NFR) from the genome of S. cerevisiae, with the use of textual representations. We employed 3 different genomic sequence representations (Hidden Markov Models, Bag-of-Words and N-gram Graphs) combined with a number of machine learning algorithms on the task of classifying genomic sequences as nucleosome-free (NFR) or nucleosome-occupied NOS (to be further amended based on updated results). We found that different approaches that involve the usage of different representations or algorithms can be more or less effective at predicting nucleosome positioning based on the textual data of the underlying genomic sequence. More interestingly, we show that N-gram Graphs, a sequence representation that takes into account both k-mer occurrences and relative positioning at various lengths scales is outperforming other methodologies and may thus be a choice of preference for the analysis of DNA sequences with subtle constraints.

Download Full-text

Single-cell chromatin interactions reveal regulatory hubs in dynamic compartmentalized domains

10.1101/307405 ◽

2018 ◽

Cited By ~ 4

Author(s):

A. Marieke Oudelaar ◽

James O.J. Davies ◽

Lars L.P. Hanssen ◽

Jelena M. Telenius ◽

Ron Schwessinger ◽

...

Keyword(s):

Single Cells ◽

Regulation Of Gene Expression ◽

Regulatory Elements ◽

Structural Basis ◽

Chromatin Domains ◽

Chromosome Conformation ◽

Chromatin Interactions ◽

Mammalian Genes ◽

Robust Regulation ◽

Order Structures

AbstractThe promoters of mammalian genes are commonly regulated by multiple distal enhancers, which physically interact within discrete chromatin domains. How such domains form and how the regulatory elements within them interact within single cells is not understood. To address this we developed Tri-C, a new Chromosome Conformation Capture (3C) approach to identify concurrent chromatin interactions at individual alleles within single cells. The heterogeneity of interactions observed between such cells shows that CTCF-mediated formation of chromatin domains and interactions within them are dynamic processes. Importantly, our analyses reveal higher-order structures involving simultaneous interactions between multiple enhancers and promoters within individual cells. This provides a structural basis for understanding how multiple cis-elements act together to establish robust regulation of gene expression.

Download Full-text

Dynamic 3D Locus Organization and Its Drivers Underpin Immunoglobulin Recombination

Frontiers in Immunology ◽

10.3389/fimmu.2020.633705 ◽

2021 ◽

Vol 11 ◽

Author(s):

Carolyn H. Rogers ◽

Olga Mielczarek ◽

Anne E. Corcoran

Keyword(s):

Large Scale ◽

New Technologies ◽

Adaptive Immune System ◽

Regulatory Elements ◽

Common Mechanism ◽

Chromosome Conformation ◽

Igh Locus ◽

Horizon Scanning ◽

V Genes ◽

Repertoire Sequencing

A functional adaptive immune system must generate enormously diverse antigen receptor (AgR) repertoires from a limited number of AgR genes, using a common mechanism, V(D)J recombination. The AgR loci are among the largest in the genome, and individual genes must overcome huge spatial and temporal challenges to co-localize with optimum variability. Our understanding of the complex mechanisms involved has increased enormously, due in part to new technologies for high resolution mapping of AgR structure and dynamic movement, underpinning mechanisms, and resulting repertoires. This review will examine these advances using the paradigm of the mouse immunoglobulin heavy chain (Igh) locus. We will discuss the key regulatory elements implicated in Igh locus structure. Recent next generation repertoire sequencing methods have shown that local chromatin state at V genes contribute to recombination efficiency. Next on the multidimensional scale, we will describe imaging studies that provided the first picture of the large-scale dynamic looping and contraction the Igh locus undergoes during recombination. We will discuss chromosome conformation capture (3C)-based technologies that have provided higher resolution pictures of Igh locus structure, including the different models that have evolved. We will consider the key transcription factors (PAX5, YY1, E2A, Ikaros), and architectural factors, CTCF and cohesin, that regulate these processes. Lastly, we will discuss a plethora of recent exciting mechanistic findings. These include Rag recombinase scanning for convergent RSS sequences within DNA loops; identification of Igh loop extrusion, and its putative role in Rag scanning; the roles of CTCF, cohesin and cohesin loading factor, WAPL therein; a new phase separation model for Igh locus compartmentalization. We will draw these together and conclude with some horizon-scanning and unresolved questions.

Download Full-text

Some comments about observations and image processing of comet 29P/Schwassmann-Wachmann 1

International Astronomical Union Colloquium ◽

10.1017/s0252921100031493 ◽

1999 ◽

Vol 173 ◽

pp. 243-248

Author(s):

D. Kubáček ◽

A. Galád ◽

A. Pravda

Keyword(s):

Image Processing ◽

Large Scale ◽

Harvard College ◽

Oak Ridge ◽

Large Scale Structures ◽

Short Period Comet ◽

Short Period ◽

Scale Structures ◽

Harvard College Observatory ◽

Period Comet

AbstractUnusual short-period comet 29P/Schwassmann-Wachmann 1 inspired many observers to explain its unpredictable outbursts. In this paper large scale structures and features from the inner part of the coma in time periods around outbursts are studied. CCD images were taken at Whipple Observatory, Mt. Hopkins, in 1989 and at Astronomical Observatory, Modra, from 1995 to 1998. Photographic plates of the comet were taken at Harvard College Observatory, Oak Ridge, from 1974 to 1982. The latter were digitized at first to apply the same techniques of image processing for optimizing the visibility of features in the coma during outbursts. Outbursts and coma structures show various shapes.

Download Full-text